Molded plastic microfluidic devices packed with self-assembled silica nanoparticles can potentially dramatically improve the resolution of the gradient elution liquid chromatographic separation of glycosylated proteins while consuming minute amount of sample. Using multiple channels (over 100) in the same device will greatly enhance throughput. This separation complements existing two-dimensional protein separations using isoelectric focusing and molecular sieving in a 2D gel. A proof of principle prototype plastic microfluidic device that is compatible with all common mobile phases of the separation methods has been fabricated. This device potentially allows the optimized packing of self-assembled nanoparticles in the channel as chromatographic column material. The Phase 1 of this proposal project focuses on developing methods for packing the channels, and surface modification of the silica particles to optimize the separation of model glycolsylated proteins using optical detection. In Phase II of the project, devices with multiple separation channels interfaced for nanospray mass spectrometry will be developed to increase throughput and detection sensitivity.